Histone Deacetylase 1 Phosphorylation Promotes Enzymatic Activity and Complex Formation

Abstract

Accessibility of the genome to DNA-binding transcription factors is regulated by proteins that control the acetylation of amino-terminal lysine residues on nucleosomal histones. Specifically, histone deacetylase (HDAC) proteins repress transcription by deacetylating histones. To date, the only known regulatory mechanism of HDAC1 function is via interaction with associated proteins. Although the control of HDAC1 function by protein interaction and recruitment is well precedented, we were interested in exploring HDAC1 regulation by post-translational modification. Human HDAC1 protein was analyzed by ion trap mass spectrometry, and two phosphorylated serine residues, Ser 421 and Ser 423 , were unambiguously identified. Loss of phosphorylation at Ser 421 and Ser 423 due to mutation to alanine or disruption of the casein kinase 2 consensus sequence directing phosphorylation reduced the enzymatic activity and complex formation of HDAC1. Deletion of the highly charged carboxyl-terminal region of HDAC1 also decreased its deacetylase activity and protein associations, revealing its requirement in maintaining HDAC1 function. Our results reinforce the importance of protein associations in modulating HDAC1 function and provide the first step toward characterizing the role of post-translational modifications in regulating HDAC activity in vivo.Although 30,000 -40,000 genes exist in a human cell, only a fraction of those genes are transcribed into mRNA and ultimately translated into the encoding protein in any given cell type (1, 2). Transcription is regulated at many levels to selectively express only those proteins necessary for proper cell function. Although the role of DNA-binding transcription factors in controlling gene expression is well established, an additional level of regulation has been recently elucidated, the accessibility of genomic DNA to transcription factors (3).The genome is tightly packed into chromosomes through nucleosomal protein-DNA complexes. Each nucleosome contains ϳ200 base pairs of double-stranded DNA wrapped twice around a histone protein core (4). Although the carboxyl-terminal region of the histones are involved in forming the nucleosome core, the amino-terminal, lysine-rich tails are located outside of the nucleosome core (5). Various studies revealed a correlation between the acetylation of amino-terminal lysine residues on histones and transcriptional activity in chromatin (3,6). Therefore, the current model of transcriptional control includes DNA accessibility due to acetylation of histones.The acetylation state of histones is dependent on the activity of two proteins, histone acetyltransferase and histone deacetylase (HDAC), 1 which acetylate and deacetylate histones, respectively. Loss of HDAC activity using small molecule inhibitors correlates with the hyperacetylation of nucleosomal histones (7,8). Importantly, HDAC proteins in yeast and humans are necessary to achieve the full transcriptional repression and induction of diverse genes (9, 10).Not surprisingly, HDAC proteins are critica...

Histone Deacetylases 1 and 2 Are Phosphorylated at Novel Sites during Varicella-Zoster Virus Infection

“…After treatment with CIAP, HDAC1 migrated more rapidly, demonstrating that it was phosphorylated by cellular kinases in uninfected cells, as previously described (38). As expected, when lysates from VZV-infected cells were examined, HDAC1 migrated as two distinct species.…”

“…As modulators of chromatin activity, HDACs are targeted during viral infection to block host gene silencing activities. It is known that HDAC activity is controlled by posttranslational modifications (8,38,49). To determine if HDAC1 and HDAC2 are modified during VZV infection, MeWo cells were either mock infected or infected with a wildtype VZV recombinant containing an amino-terminal fusion of EGFP to ORF66p.…”

“…Because of this high level of homology and their coexistence in the same protein complexes, these proteins play redundant and essential roles (33,53). Cellular proteins posttranslationally modify both HDAC1 and HDAC2, and these modifications play a role in regulating their function (8,38,49). The best characterized of these posttranslational modifications is phosphorylation by cellular kinases within the highly conserved C termini of these proteins (38,49).…”

Deactylase Inhibitors Disrupt Cellular Complexes Containing Protein Phosphatases and Deacetylases

“…HDAC6 is also phosphorylated in cells 3 and might similarly be dephosphorylated by PP1. By analogy to other HDACs, the phosphorylation of HDAC6 may regulate either its enzyme activity or its association (38,47) with other proteins as seen for HDAC11 (48) or control its cytoplasmic-nuclear shuttling as noted for HDAC7 (49) and -4 (50). HDAC6 phosphorylation could also modulate the function of the C-terminal BUZ or PAZ domain that bind ubiquitin-conjugated proteins (51) and deubiquinating enzymes and thereby regulate the ubiquitination and turnover of HDAC6 (32).…”

Regulation and Processing of Maize Histone Deacetylase Hda1 by Limited Proteolysis

“…It should be mentioned in this context that dephosphorylation of the nucleolar maize HD2 resulted in a complete loss of enzymatic activity (Kölle et al, 1999). The regulatory role of the post-translational phosphorylation of histone deacetylases in controlling both enzymatic activity and the ability to interact with other proteins was reported recently in animal cell systems (Bertos et al, 2001;Pflum et al, 2001;Tsai and Seto, 2002;Zhang et al, 2002). It is possible that phosphorylation and limited proteolysis are interrelated in the molecular response to cellular or environmental stimuli; it also is possible that ZmHda1 is enzymatically active toward nonhistone proteins, as was shown recently for HDAC6 and tubulin (Hubbert et al, 2002;Zhang et al, 2003), and then is converted to p48 with altered substrate specificity (core histones) by phosphorylation/dephosphorylation-triggered proteolysis.…”